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Liquid Handling

Xuemei Liu, Pei Hu, Yongsheng Wang, Xizhu Wang, Jinghua Huang, Jin Li, Cheng Li, Hongyun Wang, Ji Jiang
Monocyte locomotion inhibitory factor (MLIF, Met-Gln-Cys-Asn-Ser), a pentapeptide with anti-inflammatory activity, was developed for neural protection in acute ischemic stroke. Determination of MLIF in human plasma samples is of great importance for pharmacokinetic evaluation in clinical studies. A reliable and sensitive method based on ultra-performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS) was established for the measurement of MLIF in human plasma. Instability of peptide in matrix was the primary challenge in method development, which was properly resolved by addition of acidification reagents like sulfuric acid...
April 10, 2018: Journal of Pharmaceutical and Biomedical Analysis
Koray Sekeroglu, C W Extrand, David A Burdge
The onset of drainage of liquids from plastic tubes was evaluated. One end of the tubes was plugged, filled with liquid, oriented horizontally, and attached to a rotor. With their open end facing outward, the filled tubes were spun at progressively higher speeds until they began to drain. Resistance to drainage was independent of the tube length, but depended on the surface tension and density of the liquids as well as the diameter and wettability of the tubing. We found that the onset of drainage could be explained in terms of a critical centrifugal acceleration...
May 15, 2018: Langmuir: the ACS Journal of Surfaces and Colloids
Saleh Alseekh, Si Wu, Yariv Brotman, Alisdair R Fernie
Recent methodological advances in both liquid chromatography-mass spectrometry (LC-MS) and gas chromatography-mass spectrometry (GC-MS) have facilitated the profiling highly complex mixtures of primary and secondary metabolites in order to investigate a diverse range of biological questions. These techniques usually face a large number of potential sources of technical and biological variation. In this chapter we describe guidelines and normalization procedures to reduce the analytical variation, which are essential for the high-throughput evaluation of metabolic variance used in broad genetic populations which commonly entail the evaluation of hundreds or thousands of samples...
2018: Methods in Molecular Biology
Xinyuan Zhao, Wanjun Zhang, Tong Liu, Hangyan Dong, Junjie Huang, Changqing Sun, Guangshun Wang, Xiaohong Qian, Weijie Qin
Liquid biopsies using body fluids have gained much attention in recent years due to their multiple advantages in clinical diagnosis, such as less/non-invasive collection, suitability for longitudinal disease monitoring, and better representation of tumor heterogeneity. As an attractive choice for liquid biopsy, urine proteins and their post-translational modifications (PTMs) have the potential to offer significant insights into physiological variations and pathological changes in the human body. However, due to the intrinsically large variability of urine proteins and their PTMs among different individuals, there is a high demand for strategies for high-throughput analysis of a large number of samples to obtain a comprehensive view and a reliable reference interval of the urine proteome...
August 1, 2018: Talanta
B K Amos, Victoria G Pook, Seth Debolt
Chemical genetics is increasingly being employed to decode traits in plants that may be recalcitrant to traditional genetics due to gene redundancy or lethality. However, the probability of a synthetic small molecule being bioactive is low; therefore, thousands of molecules must be tested in order to find those of interest. Liquid handling robotics systems are designed to handle large numbers of samples, increasing the speed with which a chemical library can be screened in addition to minimizing/standardizing error...
April 30, 2018: Journal of Visualized Experiments: JoVE
Sabine Gogolok, Ute Köber, Steven M Pollard
Transcription activator-like effectors (TALEs) contain programmable DNA-binding domains that can be fused to various effectors to manipulate genetic sequences or transcriptional state. However, the construction of plasmids encoding the modular DNA-binding domain remains challenging due to their repetitive nature. Here, we describe methods for a simple TALE assembly reaction (STAR) that uses a 68-part plasmid library to create TALEs binding to 17 bp target sequences. Manual production of many tens of TALEs can be achieved using a simple 8 h protocol, with full length sequence-verified plasmids available within a few days...
2018: Methods in Molecular Biology
Dominique R Perez, Christian K Nickl, Anna Waller, Cristina Delgado-Martin, Travis Woods, Nitesh D Sharma, Michelle L Hermiston, Mignon L Loh, Stephen P Hunger, Stuart S Winter, Alexandre Chigaev, Bruce Edwards, Larry A Sklar, Ksenia Matlawska-Wasowska
Kinase inhibitors have dramatically increased patient survival in a multitude of cancers, including hematological malignancies. However, kinase inhibitors have not yet been integrated into current clinical trials for patients with T-cell-lineage acute lymphoblastic leukemia (T-ALL). In this study, we used a high-throughput flow cytometry (HTFC) approach to test a collection of small-molecule inhibitors, including 26 FDA-approved tyrosine kinase inhibitors in a panel of T-ALL cell lines and patient-derived xenografts...
May 1, 2018: SLAS Discovery
Jin Hui Paik, Soo Kang, Areum Durey, Ji Hye Kim, Ah Jin Kim
Nicotine is a dangerous substance extracted from tobacco leaves. When nicotine is absorbed in excessive amounts, it can lead to respiratory failure and cardiac arrest. The commercialization of electronic cigarettes (e-cigarettes) has allowed users to directly handle e-cigarette liquid. Consequently, the risk of liquid nicotine exposure has increased. We describe our experience of managing the case of a patient who orally ingested a high concentration of liquid nicotine from e-cigarette liquid. The patient presented with bradycardia and hypotension, which are symptoms of parasympathetic stimulation, together with impaired consciousness...
March 2018: Revista Brasileira de Terapia Intensiva
Annemarie Schönfeld, Mihai Constantinescu, Kirsten Peters, Martin Frenz
Electrospun meshes have been extensively investigated for tissue engineering and drug delivery. The application of this technology is of interest for laser-assisted vascular anastomosis (LAVA) due to the possibility to bind and stabilize macromolecules in fibers.
 Materials and Methods: We prepared bovine serum albumin (BSA) blend microfibers from the auxiliary proteins polyethylene oxide (PEO), polycaprolactone (PCL), polyvinyl alcohol (PVA) and gelatin. The thickness and weight of the resulting patches were measured and the morphological characteristics were observed by scanning electron microscopy...
May 9, 2018: Biomedical Materials
Phoevos Koukouvinis, George Strotos, Qingyun Zeng, Silvestre Roberto Gonzalez-Avila, Andreas Theodorakakos, Manolis Gavaises, Claus Dieter-Ohl
The present paper focuses on the simulation of the growth and collapse of a bubble in the vicinity of a wall. Both liquid and gas phases are assumed compressible and their interaction is handled with the Volume Of Fluid (VOF) method. The main interest is to quantify the influence of the induced shear stress and pressure pulse in the vicinity of the wall, for a variety of bubble sizes and bubble wall distances. Results are validated against prior experimental results, such as measurements of the bubble size, induced pressure field and shear stress on the wall...
May 8, 2018: Langmuir: the ACS Journal of Surfaces and Colloids
Sinéad Knight, Helen Plant, Lisa McWilliams, Mark Wigglesworth
Adaptation of phenotypic cell assays to 1536-well format brings major challenges in liquid handling for high-content assays requiring washing steps and coating of plates. In addition, problematic edge effects and reduced assay quality are frequently encountered. In this chapter, we describe the novel application of a centrifugal plate washer to facilitate miniaturization of 1536-well cell assays and a combination of techniques to reduce edge effects, all of which improved throughput and data quality. Cell assays currently limited in throughput because of cost and complex protocols may be enabled by the techniques presented in this chapter...
2018: Methods in Molecular Biology
Sebastian Andris, Michaela Wendeler, Xiangyang Wang, Jürgen Hubbuch
Antibody drug conjugates (ADCs) form a rapidly growing class of biopharmaceuticals which attracts a lot of attention throughout the industry due to its high potential for cancer therapy. They combine the specificity of a monoclonal antibody (mAb) and the cell-killing capacity of highly cytotoxic small molecule drugs. Site-specific conjugation approaches involve a multi-step process for covalent linkage of antibody and drug via a linker. Despite the range of parameters that have to be investigated, high-throughput methods are scarcely used so far in ADC development...
May 4, 2018: Journal of Biotechnology
Joosu Kuivanen, Sami Holmström, Birgitta Lehtinen, Merja Penttilä, Jussi Jäntti
Due to the rapidly increasing sequence information on gene variants generated by evolution and our improved abilities to engineer novel biological activities, microbial cells can be evolved for the production of a growing spectrum of compounds. For high productivity, efficient carbon channeling towards the end product is a key element. In large scale production systems the genetic modifications that ensure optimal performance cannot be dependent on plasmid-based regulators, but need to be engineered stably into the host genome...
May 5, 2018: Biotechnology Journal
Guoping Yang, Qi Pei, Chengxiao Fu, Jie Huang, Jun Chen, Shuang Yang, Yuxia Xiang, Jing Wang, Hongyi Tan, Xiaolei Liu
Pyragrel is a novel thromboxane A2-synthetase inhibitor for the treatment of cerebral infarction, and it is currently being investigated in phase I clinical trials. This paper reports the first reliable LC-MS/MS method for the simultaneous determination of Pyragrel and its two main metabolites, M1 and M2, in human plasma. All analytes were extracted from human serum using liquid-phase extraction and separated on a Zorbax EcLipse XDB C18 column using isocratic elution with a mobile phase composed of methanol, water and formic acid (65:35:0...
April 14, 2018: Journal of Pharmaceutical and Biomedical Analysis
M Adelaide Asnaghi, Ralph Duhr, Helen Quasnichka, Anthony P Hollander, Wael Kafienah, Ivan Martin, David Wendt
Bioreactor systems will likely play a key role in establishing regulatory compliant and cost-effective production systems for manufacturing engineered tissue grafts for clinical applications. However, the automation of bioreactor systems could become considerably more complex and costly due to the requirements for additional storage and liquid handling technologies if unstable supplements are added to the culture medium. Ascorbic acid (AA) is a bioactive supplement that is commonly presumed to be essential for the generation of engineered cartilage tissues...
May 4, 2018: Journal of Tissue Engineering and Regenerative Medicine
D Baunsgaard, A D Nielsen, P F Nielsen, A Henriksen, A K Kristensen, H W Bagger, M Ezban
INTRODUCTION: Advances in analytical technologies enable investigation of possible correlations between molecular structure, aggregation and subvisible particle content. Regulatory agencies place increasing attention on potential risks associated with protein aggregates in the micron range in biological therapeutics. AIM: Assess the heterogeneity, high-molecular-weight protein (HMWP) species, subvisible particle content and posttranslational modifications in six commercially available recombinant FVIII (rFVIII) products...
May 3, 2018: Haemophilia: the Official Journal of the World Federation of Hemophilia
Kyle W Anderson, Elyssia S Gallagher, Jeffrey W Hudgens
Membrane proteins are currently the most common targets for pharmaceuticals. However, characterization of their structural dynamics by hydrogen/deuterium exchange mass spectrometry (HDX-MS) is sparse due to insufficient automated methods to handle full-length membrane proteins in lipid bilayers. Additionally, membrane lipids used to mimic the membrane environment and to solubilize membrane proteins can impair chromatography performance and cause ion suppression in the mass spectrometer. The workflow discussed herein advances HDX-MS capabilities and other MS applications for membrane proteins by providing a fully automated method for HDX-MS analysis based on a phospholipid removal scheme compatible with robotic handling...
May 9, 2018: Analytical Chemistry
Evans A Monyoncho, Vlad Zamlynny, Tom K Woo, Elena A Baranova
Infrared spectroscopy is a powerful non-destructive technique for the identification and quantification of organic molecules widely used in scientific studies. For many years, efforts have been made to adopt this technique for the in situ monitoring of reactions. From these efforts, polarization modulation infrared reflection absorption spectroscopy (PM-IRRAS) was developed three decades ago. Unfortunately, because of the complexity of data processing and interpretation, PM-IRRAS had been avoided in lieu of the single potential alteration infrared spectroscopy (SPAIRS) and subtractively normalized interfacial Fourier transform infrared (SNIFTIR)...
May 3, 2018: Analyst
Brian Nguyen, Percival J Graham, Chelsea M Rochman, David Sinton
A platform compatible with microtiter plates to parallelize environmental treatments to test the complex impacts of multiple stressors, including parameters relevant to climate change and point source pollutants is developed. This platform leverages (1) the high rate of purely diffusive gas transport in aerogels to produce well-defined centimeter-scale gas concentration gradients, (2) spatial light control, and (3) established automated liquid handling. The parallel gaseous, aqueous, and light control provided by the platform is compatible with multiparameter experiments across the life sciences...
April 2018: Advanced Science (Weinheim, Baden-Wurttemberg, Germany)
Keiji Mochida
In conventional vitrification methods, the embryos are vitrified under considerable supercooling (i.e., under nonequilibrium conditions). This protocol is a refinement of newer equilibrium vitrification methods. The equilibrium vitrification solution contains a high concentration of cryoprotectants that increase its osmolality and allows vitrified embryos to survive not only during storage in liquid nitrogen (LN2 ) at -196°C but also during temporary holding at -80°C (for at least 5 mo). This high-osmolality vitrification (HOV) method is as simple as conventional vitrification and provides essentially the same high postwarming survival rate...
May 1, 2018: Cold Spring Harbor Protocols
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